Three dimensional cell cultures and light sheet-based fluorescence microscopy represent promising tools in phenotypic drug screening and personalized medicine

Undesired cytotoxic effects of many cancer drug candidates are mostly recognized quite late during the clinical trials. Just one out of ten drug candidates entering phase I trials reaches the market. This low success rate drives the high development costs and the prices of successfully marketed drugs. New assays predicting the efficacy of drugs in the early, pre-clinical stage are required to increase the success rate in drug development. Modern assays are performed with three-dimensional cell cultures. Three-dimensional cultures essentially avoid hard and flat surfaces. Hence, they favor less constrained physiological sample dynamics and promote cells growth in tissue-like environments. Conventional two-dimensional (2D) cell monolayers represent the state-of-the-art of cell-based assays in drug screening and toxicology. However, 2D systems do not reproduce the complex three-dimensional architecture of tissues. Thus, 2D cellular assays have a limited physiological significance and low predictivity to toxic compounds or drugs. Reliable screening assays based on three-dimensional cell cultures are necessary to reduce the effort and the cost of drug development and reduce failure rate of clinical studies.

We develop three-dimensional cultures of tumor cell spheroids for phenotypic drug screening issues. The cultivated spheroids resemble their microenvironment and serve as a model for tumors in the living organism (Figure 1). Tumor spheroids are easy to produce and culture. They do not need an exogenous extracellular matrix. They are well suited for high-content imaging with fluorescence microscopy and for biochemical assays. We evaluate the therapeutic benefits as well as the toxicity of drug candidates in spheroids by means of advanced light microscopy and light sheet-based fluorescence microscopy (LSFM). We develop a fluorescence imaging workstation for three-dimensional cultures based on LSFM that allows dynamic long-term observations of living three-dimensional cultures (Figure 2). Furthermore, we provide new solution to handle huge 3D data sets that arise from a time-lapse experiment, including multiple view angles and hundreds of samples by adapted data analysis algorithms and work flows. Next to cancer research, we also expect to promote novel scientific approaches in modern 3D cell biology like tissue homeostasis and development, inflammation, membrane dynamics and immunology. All the methods we employ as well as the technology we validated fulfill the requirements of good-laboratory practice.

Figure 1: Heterotypic spheroid formation of breast cancer cells (T-47D) and Normal Human Dermal Fibroblasts (NHDF).

Figure 2: Time lapse imaging of 3D cell cultures by the LSFM. (A) Perfusion system for live cell imaging in the LSFM. (B) Temperature distribution and pH stability within the perfusion system. (C) Analysis of mitotic events during spheroid formation of HepaRG cells.

Involved group members
Nariman Ansari
Francesco Pampaloni
Sabine Fischer
Biena Mathew
Alexander Schmitz
Christian Mattheyer
Roli Richa
Berit Reinhardt
Sigrun Becker
Sven Plath

University Hospital Frankfurt am Main
Georg Speyer Haus – Institute for Tumor Biology
Bayer HealthCare
Max Planck Institute of Molecular Cell Biology and Genetics
Max Planck Institute for Brain Research
SpheroTec GmbH
InSphero AG
Merz Pharmaceuticals GmbH
Carl Zeiss Microscopy GmbH


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